Modulating gene expression in Epstein-Barr virus (EBV)-Positive B cell lines with CRISPRa and CRISPRi

Liang Wei Wang; Stephen J. Trudeau; Chong Wang; Catherine Gerdt; Sizun Jiang; Bo Zhao; Benjamin E. Gewurz

doi:10.1002/cpmb.50

Modulating gene expression in Epstein-Barr virus (EBV)-Positive B cell lines with CRISPRa and CRISPRi

Liang Wei Wang, Stephen J. Trudeau, Chong Wang, Catherine Gerdt, Sizun Jiang, Bo Zhao, Benjamin E. Gewurz

Research output: Contribution to journal › Article › peer-review

5 Scopus citations

Abstract

Epstein-Barr virus (EBV) transforms small resting primary B cells into large lymphoblastoid cells which are able to grow and survive in vitro indefinitely. These cells represent a model for oncogenesis. In this unit, variants of conventional clustered regularly interspaced short palindromic repeats (CRISPR), namely the CRISPR activation (CRISPRa) and CRISPR interference (CRISPRi) methods, are discussed in the context of gene regulation at genomic DNA promoter and enhancer elements. Lymphoblastoid B cell lines (LCLs) stably expressing nuclease-deficient Cas9 (dCas9)-VP64 (Cas9 associated with CRISPRa) or dCas9-KRAB (Cas9 associated with CRISPRi) are transduced with lentivirus that encodes a single guide RNA (sgRNA) that targets a specific gene locus. The ribonucleoprotein complex formed by the dCas9 molecule and its cognate sgRNA enables sequence-specific binding at a promoter or enhancer of interest to affect the expression of genes regulated by the targeted promoter or enhancer.

Original language	English (US)
Pages (from-to)	31.13.1-31.13.18
Journal	Current Protocols in Molecular Biology
Volume	2018
DOIs	https://doi.org/10.1002/cpmb.50
State	Published - Jan 1 2018
Externally published	Yes

Bibliographical note

Funding Information:
L.W.W. is a recipient of Singapore’s Agency for Science, Technology and Re search (A*STAR) National Science Scholarship (Ph.D.). S.J. is a recipient of the Howard Hughes Medical Institute (HHMI) International Student Research Fellowship. B.Z. is funded by the National Institutes of Health (R01AI123420 and CA047006). B.E.G. is a recipient of the Burroughs Wellcome Fund Career Award in Medical Sciences. We thank John Doench for helpful discussions.

Publisher Copyright:
© 2018 by John Wiley & Sons, Inc.

Keywords

CRISPR
Enhancer
Epstein-Barr virus
Promoter
Transcription regulation

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10.1002/cpmb.50

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abstract = "Epstein-Barr virus (EBV) transforms small resting primary B cells into large lymphoblastoid cells which are able to grow and survive in vitro indefinitely. These cells represent a model for oncogenesis. In this unit, variants of conventional clustered regularly interspaced short palindromic repeats (CRISPR), namely the CRISPR activation (CRISPRa) and CRISPR interference (CRISPRi) methods, are discussed in the context of gene regulation at genomic DNA promoter and enhancer elements. Lymphoblastoid B cell lines (LCLs) stably expressing nuclease-deficient Cas9 (dCas9)-VP64 (Cas9 associated with CRISPRa) or dCas9-KRAB (Cas9 associated with CRISPRi) are transduced with lentivirus that encodes a single guide RNA (sgRNA) that targets a specific gene locus. The ribonucleoprotein complex formed by the dCas9 molecule and its cognate sgRNA enables sequence-specific binding at a promoter or enhancer of interest to affect the expression of genes regulated by the targeted promoter or enhancer.",

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AU - Zhao, Bo

AU - Gewurz, Benjamin E.

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Modulating gene expression in Epstein-Barr virus (EBV)-Positive B cell lines with CRISPRa and CRISPRi

Abstract

Bibliographical note

Keywords

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